Mechanosensitive chloride channels on the growth cones of cultured rat dorsal root ganglion neurons

Abstract

We developed an experimental system to investigate mechanosensitivity of a single neuron, using cultured rat dorsal root ganglion cells. Highly precise mechanical stimulation was applied to various sites of the cells, using a piezo-driven glass microcapillary whose movement was computer-controlled, while whole-cell patch-clamp recordings were made from the cell bodies. When the growth cones and lamillipodia from the cell soma were mechanically stimulated, inward currents were recorded at the holding potential of −60 mV. Filopodia were most sensitive to mechanical stimulation. However, when neurites or soma of dorsal root ganglion cells were stimulated in the same way, electrical responses were hardly recorded. Two types of currents varying in time-course were observed: fast type of 100–200 ms and slow type of several seconds in duration. When the membrane potential was held at around 0 mV, both types of currents were almost abolished or even reversed, and the reversal potential was estimated to be about −2.2 mV. Replacement of extracellular sodium by tetraethylammonium did not significantly change the reversal potential. In the low-chloride solution ([Cl −] o=11.7 mM), the reversal potential was about +60 mV, as expected from the Nernst equation for chloride. These inward currents were almost completely inhibited by extracellular application of chloride channel blocker, 5-nitro-2-(3-phenylpropylamino) benzoic acid (100 μM). These results indicate that the inward currents are due to activation of mechanosensitive chloride channels, preferentially located on the growth cones of cultured dorsal root ganglion neurons.